Nicotine is a drug that is widely abused in this society costing millions of dollars in health care. It is clear that the starting point of any pharmacological approach to target nicotine addiction must begin with the understanding of the mechanistic details of the action of this drug. It is known that the actions of nicotine are mediated by its ability to activate nicotinic acetylcholine receptors (nAChRs). In this proposal we ask what physiological functions of nAChRs in the brain are. One effect of activation of nAChRs is that they increase neurotransmitter release in the brain, possibly facilitating synaptic plasticity. This effect is commonly mediated by a subset of nAChRs, one containing the alpha7 subunit (a7-nAChRs), a subset that is very effective at raising intracellular calcium levels and mediating calcium-dependent signal transduction in the brain. In this proposal we examine the mechanisms underlying increased glutamate release mediated by these receptors at the mossy fiber-CA3 synapses in the hippocampus. In the previous funding period we show that nAChR activation results in a dramatic increase in frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) dependent on ER store calcium. In this proposal we investigate whether there are ER calcium stores at the mossy fiber terminals and what their properties might be using fluorescence microscopy. We then ask whether the mechanisms underlying nAChR modulation of release are specific to mEPSCs or whether they can also influence evoked release at these synapses. We will also investigate the nature and population of vesicles that are mobilized by nAChRs using styryl dyes and photoconversion followed by electron microscopy. These studies will arrive at the mechanistic details of a novel form of plasticity discovered in our lab. Lastly, we investigate the biological significance of the effect using relevant doses of nicotine and a newly developed transgenic mouse model where cholinergic fibers are labeled with GFP-tau. These studies will set the stage for the development of pharmacological tools and therapeutic strategies to combat nicotine addiction, ones that are based on realistic models of receptor physiology. PUBLIC HEALTH RELEVANCE: The abuse of nicotine costs our society millions in health care costs to combat the resultant heart disease, cancer, and other debilitating illnesses. In this project we examine how nicotine acts as a powerful modulator of brain functions, by examining its ability to usurp important signaling pathways in the brain. Our studies will form the basis for developing effective drugs to combat nicotine addiction, one that will take into account the entirety of nicotine's effects on the brain, an approach sorely lacking at present. [unreadable] [unreadable]